The present invention relates to therapeutic agents for renal diseases or organ preservatives containing a 2,3-dihydrobenzofuran derivative as an active ingredient, as well as novel 2,3-dihydrobenzofuran derivatives.
Kidney is one of organs subjected to the most serious oxidative stress in the living body. The importance of radical injury caused by active oxygen or free radicals in the development and progress mechanism of various renal diseases such as acute renal failure, drug-induced nephropathies, glomerular nephritis, diabetic nephrosis, chronic renal failure, renal transplantation has long been pointed out. In recent years, the role of lipids in cell injury has attracted particular attention (Keane W. F., Lipids and the kidney. Kidney Int., 46:910-920, 1994; Higuchi and Sanaka, xe2x80x9cRenal Diseasesxe2x80x9d, Antioxidantsxe2x80x94Free radicals and bioprotection (Niki, Shimazaki and Mino, eds.) Gakkai Shuppan Center, 223-229, 1994; Aoyagi, xe2x80x9cTherapy with Antioxidants/Scavenger, No. 3, Renal Diseasesxe2x80x9d, Therapeutics, 26:592-596, 1992). However, the effect of antioxidants, particularly lipid-peroxidation inhibitors on renal diseases has not been well explained, and any compound having a useful lipid-peroxidation inhibitory effect as a therapeutic or prophylactic agent for renal diseases or organ preservative has not been reported.
Vitamin E (xcex1-tocopherol) is a naturally occurring potent lipid-peroxidation inhibitor and its use in renal transplantation and renal ischemia models has been reported (Marubayashi, Dohi and Kawasaki, xe2x80x9cRenal maintenance and active oxygenxe2x80x9d, Kidney and Dialysis, 24:785-790, 1988; Takenaka M., Tatsukawa Y., Dohi K., Ezaki H., Matsukawa K., Kawasaki T., Transplantation, 32:137-141, 1981), but its effect is not sufficient. This is because it acts on only surfaces of membranes and lipid of the living body, but can not produce inhibitory effect against lipid-peroxidation in the interior of membranes and lipids (Niki E., Chem. Phys. Lipids, 44:227-253, 1987). Vitamin E is also expected to endogenously have an inhibitory effect against lipid-peroxidation on surfaces of membranes and lipid, because it endogenously occurs in a significantly large amount (Nakamura, xe2x80x9cAbsorption, Distribution and Excretion of Vitamin Exe2x80x9d, Vitamin Exe2x80x94Basic and Clinical Study (Igarashi, eds.), Ishiyaku Shuppan, 33-58, 1985). However, the living body does not have a sufficient protection mechanism against lipid-peroxidation in the interior of membranes and lipids, and therefore, inhibition of lipid-peroxidation in the interior of membranes and lipids seems to have an important effect for treatment and prevention of renal diseases. The effects of use of a lipid-soluble antioxidant, probucol, in various renal disease models have been reported (Modi K. S., Schreiner G. F., Purkerson M. L., J. Lab. Clin. Med., 120:310-317, 1992; Bird J. E., Milhoan K., Wilson C. B., Young S. G., Mundy C. A., Parthasarathy S., Blantz R. C., J. Clin. Invest., 81:1630-1638, 1988; Hirano T., Mamo J. C. L., Nagano S., Sugisaki T., Nephron, 58:95-100, 1991), but such simple phenolic compounds as probucol and butylated hydroxytoluene react with lipid-peroxide radicals at a lower reactivity than xcex1-tocopherol by one or more orders of magnitude (Gotoh N., Shimizu K., Komuro E., Tsuchiya J., Noguchi N., Niki E., Biochem. Biophys. Acta, 1128:147-154, 1992; Burton G. W., Ingold K. U., J. Am. Chem. Soc., 103:6472-6477, 1981) and thus have not shown sufficient protective effect for renal functions.
Thus, a potent cytoprotective agent which inhibits lipid-peroxidation that is difficult to inhibit by Vitamin E seems to be effective for the prevention and treatment of various renal diseases and maintenance of organs.
As such compounds, 4,6-di-t-butyl-2,3-dihydrobenzofuran derivatives were found to show a potent cytoprotective effect on kidney-derived cells (JP 10-72458A/98, WO97/9701729), but no report has shown that 2,3-dihydrobenzofuran derivatives having no or only one t-butyl group show such an effect.
As a result of extensive research to solve the above problems, we found that 2,3-dihydrobenzofuran derivatives having specific substituents show a potent cytoprotective effect on kidney-derived cells, and thus accomplished the present invention.
Accordingly, the present invention provides therapeutic agents for renal diseases containing a compound of general formula (1): 
wherein
R1 represents a hydrogen atom or an acyl group,
R2, R3 and R4, which may be identical or different, each represents a hydrogen atom, a lower alkyl group or a lower alkenyl group, and
R5 and R6, which may be identical or different, each represents a hydrogen atom, an optionally substituted alkyl group, an optionally substituted alkenyl group, an optionally substituted alkynyl group or an optionally substituted aryl group, or
R5 and R6 combine to form a cycloalkyl group or a saturated heterocyclic group containing one or more oxygen atoms, sulfur atoms or alkyl-substituted nitrogen atoms,
provided that R2 and R3 can not simultaneously represent a t-butyl group, or an optically active isomer or a pharmaceutically acceptable salt thereof as an active ingredient.
The present invention also provides organ preservatives containing a compound of the above general formula (1) or an optically active isomer or a pharmaceutically acceptable salt thereof as an active ingredient.
The present invention also provides compounds of general formula (1): 
wherein
R1 represents a hydrogen atom or an optionally substituted acyl group,
R2, R3 and R4, which may be identical or different, each represents a hydrogen atom, a lower alkyl group or a lower alkenyl group, provided that any one of R2 and R3 represents a t-butyl group but both of R2 and R3 can not represent a t-butyl group, and
R5 and R6, which may be identical or different, each represents a hydrogen atom, an optionally substituted alkyl group, an optionally substituted alkenyl group, an optionally substituted alkynyl group or an optionally substituted aryl group, or
R5 and R6 combine to form a cycloalkyl group or a saturated heterocyclic group containing one or more oxygen atoms, sulfur atoms or alkyl-substituted nitrogen atoms,
provided that the following cases are excluded where:
R4 represents a 2-propenyl group;
three or more of R2, R3, R4, R5 and R6 simultaneously represent a hydrogen atom;
R2 and R4 simultaneously represent a hydrogen atom and R5 and R6 simultaneously represent a methyl group; and
R3, R4, R5 and R6 simultaneously represent a methyl group,
or optically active isomers or pharmaceutically acceptable salts thereof.
In one aspect, compounds of the above general formula (1) are represented by general formula (2): 
wherein R4 represents a hydrogen atom or a t-butyl group, and R1, R3, R5 and R6 have the same meanings as defined above.
In another aspect, compounds of the above general formula (1) are represented by general formula (3): 
wherein R1, R2, R5 and R6 have the same meanings as defined above, provided that the total carbon number of R2, R5 and R6 is at least 3.
The present invention also provides compounds of general formula (1): 
wherein
R1 represents a hydrogen atom or an optionally substituted acyl group,
R2 and R4 simultaneously represent a t-butyl group, and
R3, R5 and R6 simultaneously represent a hydrogen atom,
or optically active isomers or pharmaceutically acceptable salts thereof.
The present invention also provides compounds of general formula (1): 
wherein
R1 represents a hydrogen atom or an optionally substituted acyl group,
R2, R3 and R4, which may be identical or different, each represents a hydrogen atom, a lower alkyl group or a lower alkenyl group, provided that any of R2 and R3 can not represent a t-butyl group, and
R5 and R6, which may be identical or different, each represents an optionally substituted alkyl group containing 1 to 10 carbon atoms, an optionally substituted alkenyl group containing 2 to 10 carbon atoms, an optionally substituted alkynyl group containing 2 to 10 carbon atoms or an optionally substituted aryl group containing 6 to 10 carbon atoms, or
R5 and R6 combine to form a cycloalkyl group containing 3 to 8 carbon atoms or a saturated 5-12 membered heterocyclic group containing 1 to 3 oxygen atoms, sulfur atoms or alkyl-substituted nitrogen atoms,
provided that the following cases are excluded where:
R2, R3 and R4 are independently selected from a hydrogen atom or a methyl group;
R2 and R3 each represents a hydrogen atom, a methyl group, an isopropyl group or a 2-propenyl group and R5 and R6 simultaneously represent a methyl group;
R2 represents an isopropyl group and R3 and R4 simultaneously represent a methyl group; and
R2 represents an n-butyl group and R3, R5 and R6 simultaneously represent a methyl group,
or optically active isomers or pharmaceutically acceptable salts thereof.
In the definition of substituents for compounds of general formula (1), R1 represents a hydrogen atom or an optionally substituted acyl group. Examples of the acyl group include those containing 1 to 7 carbon atoms, such as acetyl, formyl, propyonyl, trimethylacetyl and benzoyl groups. Examples of the substituted acyl group include benzyloxycarbonyl, aminoacetyl, N-methylaminoacetyl and N,N-dimethylaminoacetyl groups. However, R1 preferably represents a hydrogen atom.
R2, R3 and R4 represent a hydrogen atom, a lower alkyl group or a lower alkenyl group. The lower alkyl group means a straight or branched alkyl group containing 1 to 6 carbon atoms, such as methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl and t-butyl groups. The lower alkenyl group means a straight or branched alkenyl group containing 2 to 6 carbon atoms, such as vinyl, allyl, butenyl and pentenyl groups.
While not wishing to be bound by any particular theory, it is believed that compounds of the present invention represented by general formula (1) inhibit lipid-peroxidation within membranes and lipids in the living body to exhibit a potent cytoprotective effect on kidney-derived cells as a result of the proper hydrophobicity conferred on the benzene ring in general formula (1) by the R2, R3 and R4 groups adjacent or proximal to the hydroxyl group or acyloxy group represented by R1Oxe2x80x94. Thus, R2, R3 and R4 preferably represent a group capable of conferring proper hydrophobicity.
Therefore, one or two of R2, R3 and R4 preferably represent an alkyl or alkenyl group containing 3 to 6 carbon atoms, more preferably a branched alkyl group containing 3 to 4 carbon atoms, and most preferably a t-butyl group.
In a preferred embodiment, one or two of R2, R3 and R4 represent(s) a branched alkyl or alkenyl group containing 3 to 6 carbon atoms, and the other(s) represent(s) a hydrogen atom. In a more preferred embodiment, one or both of R2 and R3 represent a branched alkyl or alkenyl group containing 3 to 6 carbon atoms, the total carbon number of R2 and R3 is 8 or less, and R4 represents a hydrogen atom. In a most preferred embodiment, one of R2 and R3 represents a t-butyl group, the total carbon number of R2 and R3 is 8 or less, and R4 represents a hydrogen atom. In another preferred embodiment, one or two of R2, R3 and R4 represent(s) a t-butyl group, the other(s) represent(s) a hydrogen atom, and the total carbon number of R2, R3 and R4 is 8 or less.
R5 and R6 may be identical or different and each represents a hydrogen atom, an optionally substituted alkyl group, an optionally substituted alkenyl group, an optionally substituted alkynyl group or an optionally substituted aryl group. This alkyl group means a straight or branched alkyl group containing 1 to 20, preferably 1 to 10, more preferably 1 to 6, and most preferably 2 to 6 carbon atoms, such as methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, pentyl, hexyl, heptyl, octyl, nonyl, or decyl group. The alkenyl group means a straight or branched alkenyl group containing 2 to 20, preferably 2 to 10, and more preferably 2 to 6 carbon atoms, such as vinyl, allyl, butenyl, pentenyl, geranyl, or farnesyl group. The alkynyl group means a straight or branched alkynyl group containing 2 to 20, preferably 2 to 10, and more preferably 2 to 6 carbon atoms, such as ethynyl, propynyl, or butynyl group. The aryl groups means a monovalent aromatic hydrocarbon preferably containing 6 to 20, and more preferably 6 to 10 carbon atoms, such as phenyl, tolyl, xylyl, biphenyl, naphthyl, anthryl, or phenanthryl group.
When R5 and R6 represent an alkyl, alkenyl, alkynyl or aryl group, they may have a substituent such as a halogen atom, a lower alkyl group, a lower alkenyl group, a hydroxyl group, an amino group, a substituted amino group such as dimethylamino, an alkoxy group, an aryloxy group, a nitro group, a trifluoromethyl group, a phenyl group, an acetoxy group, etc. Examples of the halogen atom include chlorine, bromine, fluorine and iodine. Examples of the lower alkyl and lower alkenyl groups include those listed above for R2. Examples of the alkoxy and aryloxy groups include those derived from the alkyl and aryl groups listed above for R5 and R6.
Alternatively, R5 and R6 combine to form a cycloalkyl group or a saturated heterocyclic group containing one or more oxygen atoms, sulfur atoms or alkyl-substituted nitrogen atoms. The cycloalkyl group here means a cycloalkyl group containing 3 to 8, preferably 5 to 8 carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, or cyclooctyl group. Examples of the saturated heterocyclic group containing oxygen atoms, sulfur atoms or alkyl-substituted nitrogen atoms include saturated 5-12 membered heterocyclic groups containing 1 to 3 such heteroatoms, such as tetrahydropyranyl, tetrahydrothiopyranyl, and N-methylpiperidyl groups.
Compounds of general formula (1) wherein R2 represents a t-butyl group preferably have the following substituents.
R1 is preferably a hydrogen atom or an acetyl, trimethylacetyl, benzyloxycarbonyl, aminoacetyl, N-methylaminoacetyl or N,N-dimethylaminoacetyl group, especially a hydrogen atom or an acetyl, trimethylacetyl or N,N-dimethylaminoacetyl group.
R3 is preferably a hydrogen atom or a methyl, ethyl, n-propyl, i-propyl or i-butyl group, especially a hydrogen atom or an i-propyl group.
R4 is preferably a hydrogen atom or a methyl, n-propyl, i-butyl or t-butyl group, especially a hydrogen atom or a t-butyl group.
R5 is preferably a hydrogen atom or a methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, n-pentyl, benzyl, hydroxymethyl, aminomethyl or N,N-dimethylaminomethyl group, especially a hydrogen atom or a methyl, n-pentyl, benzyl, hydroxymethyl, aminomethyl or N,N-dimethylaminomethyl group.
R6 is preferably a hydrogen atom or a methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, n-pentyl or benzyl group, especially a hydrogen atom or a methyl, n-pentyl or benzyl group.
The cyclic group formed by R5 and R6 is preferably a cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, tetrahydropyranyl, tetrahydrothiopyranyl or N-methylpiperidyl group, especially a cyclopentyl, cyclohexyl, cycloheptyl or tetrahydropyranyl group.
Compounds of general formula (1) wherein R3 represents a t-butyl group preferably have the following substituents.
R1 is preferably a hydrogen atom or an acetyl, trimethylacetyl, benzyloxycarbonyl, aminoacetyl, N-methylaminoacetyl or N,N-dimethylaminoacetyl group, especially a hydrogen atom or an acetyl, trimethylacetyl or N,N-dimethylaminoacetyl group.
R2 is preferably a hydrogen atom or a methyl, ethyl, n-propyl, i-propyl, i-butyl, s-butyl, allyl or methallyl group, especially a hydrogen atom or a methyl, n-propyl, i-propyl, i-butyl or s-butyl group.
R4 is preferably a hydrogen atom or a methyl or n-propyl group, especially a hydrogen atom.
R5 is preferably a hydrogen atom or a methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, n-pentyl, benzyl, hydroxymethyl, aminomethyl or N,N-dimethylaminomethyl group, especially a hydrogen atom or a methyl, n-pentyl, benzyl, hydroxymethyl, aminomethyl or N,N-dimethylaminomethyl group.
R6 is preferably a hydrogen atom or a methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, s-butyl, t-butyl, n-pentyl or benzyl group, especially a hydrogen atom or a methyl, n-pentyl or benzyl group.
The cyclic group formed by R5 and R6 is preferably a cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, tetrahydropyranyl, tetrahydrothiopyranyl or N-methylpiperidyl group, especially a cyclopentyl, cyclohexyl, cycloheptyl or tetrahydropyranyl group.
In a preferred embodiment, R1 represents a hydrogen atom or an acyl group containing 1 to 7 carbon atoms; one or two of R2, R3 and R4 represent(s) a branched alkyl or alkenyl group containing 3 to 6 carbon atoms while the other(s) represent(s) a hydrogen atom; and R5 and R6 each represents a hydrogen atom or an alkyl group containing 1 to 20 carbon atoms or R5 and R6 combine to form a cycloalkyl group containing 3 to 8 carbon atoms. In a more preferred embodiment, R1 represents a hydrogen atom; one or both of R2 and R3 each represents a branched alkyl or alkenyl group containing 3 to 6 carbon atoms but the total carbon number of R2 and R3 is 8 or less; R4 represents a hydrogen atom; and R5 and R6 each represents a hydrogen atom or an alkyl group containing 1 to 1 carbon atoms or R5 and R6 combine to form a cycloalkyl group containing 3 to 8 carbon atoms. In a most preferred embodiment, R1 represents a hydrogen atom; one of R2 and R3 represents a t-butyl group but the total carbon number of R2 and R3 is 8 or less; R4 represents a hydrogen atom; and R5 and R6 each represents a hydrogen atom or an alkyl group containing 1 to 6 carbon atoms or R5 and R6 combine to form a cycloalkyl group containing 5 to 8 carbon atoms. In another preferred embodiment, R1 represents a hydrogen atom; one or two of R2, R3 and R4 represent(s) a t-butyl group while the other(s) represent(s) a hydrogen atom but the total carbon number of R2, R3 and R4 is 8 or less; and R5 and R6 each represents a hydrogen atom or an alkyl group containing 1 to 6 carbon atoms or R5 and R6 combine to form a cycloalkyl group containing 5 to 8 carbon atoms.
Compounds of the present invention represented by general formula (1) may have an asymmetric center, therefore may be optically active. The present invention includes not only racemates but also optically active isomers themselves.
Compounds of general formula (1) having some substituents can form an addition salt with an acid or a base. Therefore, the present invention includes pharmaceutically acceptable salts of compounds of general formula (1). Examples of the acid addition salt of the compound of general formula (1) include inorganic salts such as hydrochloride, sulfate, nitrate and phosphate, as well as organic acid salts such as acetate, lactate, oxalate, citrate, tartrate and p-toluenesulfonate. Examples of the base addition salt of the compound of general formula (1) include salts with inorganic bases such as sodium salt, potassium salt, calcium salt, aluminium salt, ammonium salt, as well as organic amine salts.
Preferred examples of the compounds of the present invention are as follows:
4,7-di-t-butyl-5-hydroxy-2,3-dihydrobenzofuran;
4,7-di-t-butyl-5-hydroxy-2-methyl-2,3-dihydrobenzofuran;
4,7-di-t-butyl-5-hydroxy-2-n-octyl-2,3-dihydrobenzofuran;
4,7-di-t-butyl-5-hydroxy-2,2-dimethyl-2,3-dihydrobenzofuran;
6-t-butyl-5-hydroxy-2,2-dimethyl-2,3-dihydrobenzofuran;
6-t-butyl-2,2-di-n-pentyl-5-hydroxy-2,3-dihydrobenzofuran;
6-t-butyl-5-hydroxy-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-(2-propenyl)-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-n-propyl-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-(2-methyl-2-propenyl)-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-(2-methylpropyl)-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-(1-methyl-2-propenyl)-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-(1-methylpropyl)-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-4-methyl-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-4-ethyl-5-hydroxy-2,3-dihydrobenzofuran-2-spiro-1xe2x80x2-cycloheptane;
6-t-butyl-5-hydroxy-2,3-dihydrobenzofuran;
6-t-butyl-5-hydroxy-2-methyl-2,3-dihydrobenzofuran;
2,2-diethyl-5-hydroxy-4,6-diisopropyl-2,3-dihydrobenzofuran;
5-hydroxy-2,2-dimethyl-4,6-diisopropyl-2,3-dihydrobenzofuran;
6-t-butyl-5-hydroxy-2,2,4-trimethyl-2,3-dihydrobenzofuran;
6-t-butyl-5-hydroxy-2,2-dimethyl-4-isopropyl-2,3-dihydrobenzofuran; and
4-t-butyl-5-hydroxy-2,2-dimethyl-6-isopropyl-2,3-dihydrobenzofuran.
Therapeutic agents for renal diseases of the present invention can be used as various pharmaceutical compositions comprising a compound of general formula (1) as an active ingredient in combination with a physiologically non-toxic solid or liquid pharmaceutical carrier. These pharmaceutical compositions are formulated and used in various dosage forms depending on the administration route. Dosage forms include tablets, granules, pills, capsules, solutions, syrups, suspensions, emulsions and injections. Suitable pharmaceutical carriers include commonly used excipients, binders, disintegrants, lubricants, coating agents, dissolution-aids, emulsifiers, suspending agents, stabilizers and solvents.
Therapeutic agents for renal diseases of the present invention can be used via oral administration, parenteral administration such as intravenous injection, administration of sustained-release formulations, etc.
Therapeutic agents for renal diseases of the present invention can be used for therapies of various renal diseases such as chronic renal failure, diabetic nephrosis, glomerular nephritis, immunocomplex nephritis, acute renal failure, nephropathies caused by platinum complex-based anticancer agents such as cisplatin or other drugs such as gentamicin, nephropathies caused by agrichemicals such as Paracort, and uremia. Actually desired dosage of compounds of general formula (1) depends on the age, sex and weight of the patient, severity of condition, administration route or other factors, and an effective daily dosage which is normally acceptable is, for example, 1-1000 mg, and preferably 10-500 mg/adult. Such dosage is preferably divided into 1-3 doses per day per patient in need of therapy.
Compounds of general formula (1) can also be used as an active ingredient of organ preservatives. Compounds of general formula (1) can be used as organ preservatives for any organ of humans and animals, such as brain, heart, kidney, pancreas, lung, liver and bone marrow cells, preferably kidney of humans and animals. Compounds of the present invention can be added in a maintenance solution or a perfusion solution to minimize damages of an organ during storage of the organ extracted from a donor for transplantation. Compounds of the present invention can be used to inhibit deterioration of extracted organs and to maintain functions of the organs after transplantation.
When compounds of general formula (1) are used in a maintenance solution required for storing organs, they are preferably dissolved at an effective dose which is normally acceptable such as 1-1000 mg in the maintenance solution at a concentration of, for example, 0.1-10000 mg/L.
The following examples and test examples further illustrate the present invention, but are not construed as limiting the same.